Method for diagnosis of monoclonality in leukaemia and lymphoma

ABSTRACT

The present invention is directed to a method for the detection of leukemia or lymphoma by determining the homogeneity or heterogeneity of the length of immunoglobulin or T-receptor gene segments. The present invention is further directed to a kit which is useful for the detection of leukemia or lymphoma.

This invention relates to a diagnostic method which may be used inleukaemia and lymphoma for detection of malignancy and determination ofits lineage. Lymphocytes, the cells responsible for immunity, are of twotypes:

(a) the B-lymphocytes, each of which produces a specific immmunoglobulinmolecule which attaches to a particular foreign antigen; and

(b) the T-lymphocytes, each of which has a specific surface receptorwhich enables the cell to attach to a particular foreign antigen.

Each B-lymphocyte contains a unique immunoglobulin gene which differsmoderately in structure from the immunoglobulin gene of all otherB-lymphocytes and markedly in structure from the immunoglobulin genes ofall other body cells, including T-lymphocytes. Similarly, eachT-lymphocyte contains a unique T-receptor gene which differs moderatelyin structure from the T-receptor gene of all other T-lymphocytes andmarkedly in structure from the T-receptor gene of all body cells,including B-lymphocytes.

Lymphoid leukaemias and lymphomas are a form of cancer of the lymphocytetissue. Each leukaemia or lymphoma arises from a single B- orT-lymphocyte which multiples, spreads and eventually results in deathunless treated. As all the tumor cells are descended from a single cell,they are all genetically the same and all will contain the same uniquegene, a unique immunoglobulin gene if the tumour arose in aB-lymphocyte, or a unique T-receptor gene if the tumour arose in aT-lymphocyte.

Conversely, if a tissue is suspected as being involved by leukaemia orlymphoma, the detection of a unique immunoglobulin gene is presumptiveevidence of a tumour of B-lymphocytes whereas detection of a uniqueT-receptor gene is presumptive evidence of a tumour of T-lymphocytes.

The object of the present invention is to determine whether or notleukaemia or lymphoma is present in a tissue sample by determiningwhether or not a monoclonal B- or T-lymphocyte population is present inthe sample. In general terms, the invention does this by focussing on adiscrete segment of the immunoglobulin or T-receptor molecule anddetermining whether all or most of these segments in the tissue sampleare rearranged and have precisely the same length, implying that theyare derived from the same unique molecule.

Present methods for detection of monoclonality are based on restrictionenzyme digestion, followed by Southern blotting and gene probing. Thisapproach is complex, expensive and time consuming, so that informationis usually provided too late to be of substantial practical value.

By contrast, the method of the present invention provides a rapid andsensitive diagnostic test. In addition, it is also very versatile asmaterial which can be used as the tissue sample includes blood, tumourtissue in node or bone marrow, formalin-fixed embedded histologicalmaterial, aspirated cytological material or cells on slides.

According to the present invention, there is provided a method forlymphoid leukaemia and/or lymphoma in a tissue sample, which comprisesthe determination of the homogeneity or heterogeneity of the length ofimmunoglobulin and/or T-receptor gene segments in said tissue sample toindicate the presence or absence of monoclonality of the B- and/orT-lymphocyte population in said sample.

According to one embodiment of the present invention there is provided amethod for the detection of monoclonality and presumptive malignancy ina tissue sample, and/or for the determination of the B-lymphocyte orT-lymphocyte origin of a tumour, which comprises the steps of:

(a) amplification of immunoglobulin and/or T-receptor gene segments insaid tissue sample by means of the polymerase chain reaction usingspecific primers or mixtures of primers for said immunoglobulin and/orT-receptor gene segments; and

(b) size separation of the amplified segments to determine homogeneityor heterogeneity of the length of the amplified segments.

In one particular embodiment, the primers used are consensus primers forthe immunoglobulin and/or T-receptor gene segments.

The present invention also extends to a kit for performance of themethod of the invention as broadly outlined above, comprising a set ofspecific primers or mixture(is of primers for immunoglobulin and/orT-receptor gene segments for amplification of immunoglobulin and/orT-receptor gene segments in a tissue sample by the polymerase chainreaction.

Once again, in a particular embodiment, the primers used are consensusprimers for the immunoglobulin and/or T-receptor gene segments.

The immunoglobulin (Ig) and T-receptor (Tr) genes are present in allcells. Each consists of 4 families, the variable (V), diversity (D),joining (i), and constant (C) region family (except for immunoglobulinlight chains which lack a D segment). In the process of development of aB- or T-lymphocyte, the gene is rearranged so that one randomly chosenmember of each family is joined together to form the final molecule.Random mutations also occur at the VD, DJ and JC joining points. As aresult of this random joining and mutation, the final immunoglobulin orT-receptor molecule is virtually unique. However, there are two featuresof particular importance with regard to the present invention:

(a) The number of bases removed and/or inserted at the VD, DJ and JCjunctions is quite variable, so that different immunoglobulin orT-receptor molecules, particularly the segments spanning VD, DJ and JCjunctions, differ substantially in length.

(b) There are some regions of similarity if not of absolute identitywithin the genes. These comprise certain parts of the V regions, termed"framework" regions, and parts of the J and C regions.

The present invention involves the use of the polymerase chain reaction(PCR) in a new way. The PCR is now well known, having been firstdescribed in 1985, and it enables the exponential amplification of smallCDNA segments provided that their sequence is known. The principle ofthe PCR is that two DNA primers, one for each DNA strand, are used insuccessive cycles of denaturation followed by DNA synthesis and thisresults in exponential amplification of the segment of DNA bounded bythe primers. As each cycle takes 6-7 minutes, it is practical to perform20-30 cycles, and as each cycle may be up to 70-80% efficient, there canbe amplification of up to 10⁶ -fold at high fidelity.

The PCR as described is based on primers, each exactly complementary toa known sequence. In one embodiment of the present invention, theproblem of amplifying immunoglobulin and T-receptor molecules has beensolved by utilizing consensus primers to regions which have a similarbut not identical sequence in the immunoglobulin and T-receptor genesrespectively. These regions comprise the framework portions of the Vregions of the immunoglobulins, conserved V regions of the T-receptorgenes, and parts of the D,J and/or C regions of the immunoglobulin orT-receptor genes. Primers to these regions work for two reasons.Firstly, they have a structure which is sufficiently homologous to theappropriate region of the immunoglobulin or T-receptor DNA or RNA torecognise only those regions in total DNA or RNA. Secondly, the PCRregion will only work if the sequence recognised by the two primers isshort. This condition only obtains for those segments of DNA or RNA inwhich the V, D and J (or C) regions have been joined together and amature immunoglobulin or T-receptor molecule has been formed. As aresult of these two properties, the primers will recognise and amplifyonly the final mature immunoglobulin or T-receptor molecule.

Use of these primers in the PCR will result in amplification of theinter-primer segments of all mature immunoglobulin or T-receptormolecules in a tissue sample. The amplified segments will cross the VD,DJ (and JC) junctions. As a result the final amplified piece of DNA,irrespective of its sequence, will have a single length if all of theimmunoglobulin or T-receptor genes in the tissue are derived from amonoclonal, malignant population or will have a heterogenous length ifthe molecules are derived from a heterogenous non-malignant population.Primers for the immunoglobulin molecule will identify malignantpopulations of B-lymphocyte origin, primers for the T-receptor moleculewill identify malignancy of T-lymphocyte origin.

The lengths of the amplified pieces of DNA can be simply determined byseparating the DNA molecules by a technique which separates molecules onthe basis of size. At present electrophoresis in agarose orpolyacrylamide gel is used but chromatography would have advantagesowing to the feasibility of automation. The size separated molecules canbe identified by non-isotopic means, e.g. ethidium bromide staining, byradioisotope incorporation, or by Southern transfer and hybridization toan internal probe.

Further features of the present invention will be apparent from thefollowing detailed description.

A. SYNTHESIS OF PRIMERS 1. Primers for the Immunoglobulin Gene

The 4 regions of DNA sequence which are similar in all immunoglobulinheavy chain genes are:

(a) in the 3' end of the J-region, 30 of 34 bases are conserved in allsix germ-line i-chains. 3 DNA primers which will stick to the (+) strandDNA have been used:

    __________________________________________________________________________    ELJ.sub.H -5'TGAGG AGACG GTGAC CAGGG TNCCT TGGCC CCAG3'                       LJ.sub.H - 5'TGAGG AGACG GTGAC C                                                                      3'                                                    VLJ.sub.H -5'                                                                            GTGAC CAGGG TNCCT TGGCC CCAG3'                                     __________________________________________________________________________

(wherein N represents any of the four nucleotide bases.)

(b)-(d) the V-region can be subdivided into 3 "framework" regions andintervening "chain determining" regions. The "framework" regions aresimilar in all immunoglobulins; the "chain determining" regions varygreatly. By comparing sequence of 17 lymphocyte lines, conservedstretches of DNA have been identified in each framework region. However,the sequences are not highly conserved. As it is not clear how mis-matchwill affect the PCR, redundancies have been introduced into somesynthetic primers. Primers which will hybridize to (-) strand of DNAinclude:

    __________________________________________________________________________    FR1BR:                                                                             5'CT[C/G] [T/A]C                                                                        CTG[T/C] [G/A]CAG[T/C]C                                                                        TCTGG 3'                                      FR1BS:                                                                             5'CTCTC   CTGTG  CAGCC     TCTGG 3'                                      FR2A:                                                                              5'TGG[A/G]T                                                                             CCG[C/A]C                                                                            AG[G/C]C[T/C]                                                                           [T/C]CNGG 3'                                  FR3A:                                                                              5'ACACG   GC[C/T] [G/C]T GTATT                                                                           ACTGT 3'                                      Primers which stick to the (+) strand of DNA include:                         FR3A-:                                                                             5'ACAGT   AATAC  A[G/C] [A/G]GC                                                                          CGTGT 3'                                      __________________________________________________________________________

Primers which stick to the opposite strand of DNA and/or for the otherimmunoglobulin chains could equally well be used.

Primers modified to contain internal or terminal restriction enzymesites have also been used successfully. They aid subsequent cloning.

2. Primers for the T-receptor Gene

Analogous primers are used for the T-receptor genes. The principlesapplied in selecting primers for these genes are the same as for theimmunoglobulin gene but there are some differences of emphasis. Inparticular, there is more emphasis on generation of diversity by removalof nucleotides and insertion of random nucleotides at VD, DJ and JCjunctions. The V-regions of the T-receptor cannot be divided easily intoless variable and "framework" and more variable "chain determining"regions. However, near the VD junction a stretch of about 20 bases isconserved.

    ______________________________________                                        JAT1   5' TTC CAA AGA TCA G(T/C)T TG 3'                                       JAT2   5' A(A/G)C CTG GT(T/C) CCT (T/G)(T/G)T                                        CAA A3'                                                                3VTA   5' GA(C/T) TCA GC(T/C) GTG TAC                                                (T/C)(T/A)(C/T) TG 3'                                                  5VTA1  5' (A/T)AC (C/T)T(A/c) (C/T)TC TGG TA(C/T)                                    AA(A/G) CA 3'                                                          CTA    5' GAA TAG GCA GAC AGA CTT GT 3'                                       PTA    5' ACT GGA TTT AGA GTC TCT 3'                                          ______________________________________                                    

JAT2 is the i-region primer for the PCR.

JAT1 is an internal probe.

3VTA is the V-region primer for the PCR, about 80 base pairs from JAT1.

5VTA is a V-region primer 200-300 base pairs from JAT1.

CTA and PTA are within the constant region.

The V-and J-region primers are consensus primers.

(b) T-receptor beta chain

    ______________________________________                                        JBT1  5' CG GGT (G/C)CC T(T/G)G CCC (G/A)AA 3'                                JBT2  5' AG CAC GGT GAG CC(G/T) (G/T)GT                                             (G/C)CC 3'                                                              3VTB1 5' GAC (T/A)(C/G)A (G/A)(G/C)C GTG TAT                                        CT(C/T) TG3'                                                            3VTB2 5' CAG ACA TCT CTG TAC (C/T)TC TGT GCC 3'                               5VTB1 5' C(A/T)(C/A)(T/C)(A/C) T (T/G)T(A/T)(T/C)                                   TGGTA CCGAC AG 3'                                                       5VTB2 5' TGTAC TGGTA TCGAC AAG 3'                                             CTB   5' TTT GGG TGT GGG AGA TCT CTG C 3'                                     PTB   5' CTT CTG ATG GCT CAA ACA 3'                                           ______________________________________                                    

JBT2 is a J-region primer for the PCR.

JBT1 is an internal probe.

3VTB1 and 3VTB2 are V-region primers about 80 base pairs from JBT1, forTCR beta subclasses Beta, and Beta2 respectively.

5VTB1 and 5VTB2 are V-region primers 200-300 base pairs from JBT1, forTCR beta subclasses Beta₁ and Beta₂ respectively.

CTB and PTB are within the constant region for all Beta genes.

The V- and J-region primers are consensus primers.

    ______________________________________                                        JGT12-    5' AAG TGT TGT TCC ACT GCC AAA 3'                                   3VTG      5' GTC TAT TAC TGT GCC ACC TGG 3'                                   CTG       5' TCT GGA GCT TTG TTT CAG CAA 3'                                   PTG       5' GGA AGA AAA ATA GTG GGC 3'                                       ______________________________________                                    

(c) T-receptor gamma chain

JGT12 is a J-region primer for the PCR. It is based on the sequence oftwo of the four known J-chains of TCR class Gamma.

3VTG is a V-region primer about 80 base pairs from JGT12. CTG and PTGare within the constant region.

As constant region primers are sometimes used for amplification,depending on the individual case, it may be necessary to precede the PCRamplification by starting from RNA and carrying out one round of reversetranscription.

The internal probes JAT1 and JBT1 described above can be used for finaldetection of an amplified product if it cannot be visualized as anethidium--stained band. The probe can be labelled, usuallyradioactively, and thereby provides a very sensitive and specific methodfor detection of the amplified product.

An alternative approach applicable to the gamma chain (Tg receptor) isto use a mixture of primers for the 9 V-regions and 5 J-regions.Sequences used are:

    __________________________________________________________________________    V-regions                                                                     Primer  Sequence                                                              __________________________________________________________________________    TCRVG2PST                                                                             5' CTTC CTGCAG ATG ACT CCT ACA ACT CCA AGG                                    TTG 3'                                                                TCRVG3PST                                                                             5' CTTC CTG CAG ATG ACG TCT CCA CCG CAA GGG                                   ATG 3'                                                                TCRVG4PST                                                                             5' CTTC CTG CAG ATG ACT CCT ACA CCT CCA GCG                                   TTG 3'                                                                TCRVG5PST                                                                             5' TTC CTG CAG ATG ACG TCT CCAA CTC AAA GGA                                   TG 3'                                                                 TCRVG8PST                                                                             5' CTTC CTGCAG ATG ACT CCT ACA ACT CCA GGG                                    TTC 3'                                                                TCRVG9PST                                                                             5' GG(A/G/C/T) ACTG CAG GAA AG GAA TCTG GCATT                                 CCG 3'                                                                TCRVG10PST                                                                            5' CT CTG CAG AAT CCG CAG CTC GAC GCA GCA 3'                          TCRVG11PST                                                                            5' CA CTG CAG GCT CAA GAT TGC TCA GGT GGG 3'                          TCRVG12PST                                                                            5' ACT CTG CAG CCT CTT GGG CAC TGC TCT AAA 3'                         __________________________________________________________________________

    ______________________________________                                        J-regions                                                                     Primer  Sequence                                                              ______________________________________                                        JGT.sub.12.spsb.-                                                                     5' AAG TGT TGT TCC ACT GCC AAA 3'                                     JGT.sub.3                                                                             5' AGTTA CTA TG AG C (T/C) T AGT CCC 3'                               JGT.sub.4                                                                             5' TGT AAT GAT AAG CTT TGT TCC 3'                                     ______________________________________                                    

The V-region primers contain 20-24 bases (those nearest the 3' end)which match the target precisely. They also contain, near the 5' end,the sequence CTGCAG, as site for a restriction enzyme. This site willaid subsequent cloning, should cloning be necessary.

B. AMPLIFICATION OF DNA

Both DNA or RNA may be used. CDNA is made from RNA using reversetranscriptase and one or both PCR primers. The CDNA is precipitated andresuspended in water and the PCR reaction then used. Approximately 30cycles of the reaction are performed using polymerase from Thermusaquaticus. Products of the reaction are separated on agarose gels andbecome visible following staining with ethidium bromide. Southernblotting and hybridization to a DNA probe which binds internal to theprimer binding sites is also usually performed. Control size-markers aidin determining size of the amplified segment.

C. RESULTS OF TESTING FOR MONOCLONALITY USING CONSENSUS IMMNOGLOBULINPRIMERS

Tables 1 and 2 set out the results of testing various samples formonoclonality using the immunoglobulin gene only.

                  TABLE 1                                                         ______________________________________                                        Results on Extracted DNA                                                      Material       Number   Result                                                ______________________________________                                        Normal T cell clones                                                                         15       no amplification                                      Comment: As predicted.                                                        Normal B cell clones                                                                         16       1 or 2 discrete bands                                 Comment: As predicted.                                                        Normal mixed blood                                                                           20       diffuse peak                                          lymphocytes                                                                   Comment: As predicted.                                                        B-cell lymphomas                                                                             24       1 or 2 discrete bands in                                                      19*, no amplification in 5**                          ______________________________________                                         Comment: *As predicted. **The cases showing no amplification probably         represent cases with an unusual form of rearrangement.                   

                  TABLE 2                                                         ______________________________________                                        Results on Fixed Material and Node Aspirates                                  Material   Number      Results                                                ______________________________________                                        Fixed material                                                                B cell lymphomas                                                                         26          1 or 2 discrete bands in 24.                           T cell lymphomas                                                                         7           no amplification                                       Reactive nodes                                                                           9           no amplification                                       Carcinomas 12          no amplification                                       Node aspirates                                                                B cell lymphomas                                                                         5           1 or 2 discrete bands in 3                             Carcinomas 2           no amplification.                                      ______________________________________                                    

These results were obtained using primers LJ_(H) ⁻ and FR3A describedabove. Thirty cycles of amplification were performed and the amplifiedfragments were electrophoresed in 2% agar and stained with ethidiumbromide to enable visualisation with ultraviolet light. Similar resultshave been obtained by adding radioactive nucleotide (CTP) for the lastfive cycles of amplification, electrophoresing, and visualising thefragments by autoradiography.

D. RESULTS OF TESTING FOR MONOCLONALITY USING T-RECEPTOR PRIMERS.

Using the same PCR procedures as described above with one or other pairsof consensus primers, a discrete rearrangement has been detected usingone or other pair of primers in 6 of 8 normal T-cell clones.

                  TABLE 3                                                         ______________________________________                                        Results on 8 normal T-cell clones using                                       consenus primers.                                                                          Number of clones showing a                                       Primer combination                                                                         discrete band                                                    ______________________________________                                        3VTA & JAT2  3                                                                3VTA & JBT2  5                                                                3VTB2 & JBT2 2                                                                5VTB1 & JBT1 5                                                                5VTB2 & JBT2 4                                                                ______________________________________                                    

Similarly, using a mixture of all 9 V-region primers as previouslydescribed (TCRVG2PST-TCRVG12PST, and JGT₁₂₋, JGT₃ and JGT₄ a discreterearrangement has been detecter in 3 of 4 normal T-cell clones.

We claim:
 1. A method for the detection of B-cell lymphoid leukemia orB-cell lymphoma in a tissue sample which comprises:(a) amplifying theimmunoglobulin gene segments in said tissue sample by the polymerasechain reaction using at least one pair of primers specific for saidimmunoglobulin gene segments, wherein said immunoglobulin gene segmentscomprise at least one of a V-D, D-J and J-C junction; (b) separating theamplified immunoglobulin gene segments by size; (c) detecting saidsize-separated amplified immunoglobulin gene fragments; and (d)determining the homogeneity or heterogeneity of said amplifiedimmunoglobulin gene segments wherein homogeneity is indicative of B-celllymphoid leukemia or B-cell lymphoma.
 2. A method for the detection ofT-cell lymphoid leukemia or T-cell lymphoma in a tissue sample whichcomprises:(a) amplifying the T-receptor gene segments in said tissuesample by the polymerase chain reaction using at least one pair ofprimers specific for said T-receptor gene segments, wherein saidT-receptor gene segments comprise at least one of a V-D, D-J and J-Cjunction; (b) separating the amplified T-receptor gene segments by size;(c) detecting said size-separated amplified T-receptor gene fragments;and (d) determining the homogeneity or heterogeneity of said amplifiedT-receptor gene segments wherein homogeneity is indicative of T-celllymphoid leukemia or T-cell lymphoma.
 3. A method for determining theT-cell or B-cell origin of a lymphoma or leukemia in a tissue samplewhich comprises:(a) amplifying the immunoglobulin and T-receptor genesegments in said tissue sample by the polymerase chain reaction using atleast one pair of primers specific for said immunoglobulin gene segmentand at least one pair of primers specific for said T-receptor genesegments, wherein said immunoglobulin gene segments and T-receptor genesegments each comprise at least one of a V-D, D-J and J-C junction; (b)separating the amplified immunoglobulin and T-receptor gene segments bysize; (c) detecting said size-separated amplified immunoglobulin andT-receptor gene fragments; and (d) determining the homogeneity orheterogeneity of said amplified T-receptor immunoglobulin gene segmentswherein homogeneity of T-receptor gene segments is indicative of T-cellorigin of said lymphoma or leukemia and homogeneity of immunoglobulingene segments is indicative of B-cell origin of said lymphoma orleukemia.
 4. The method of any one of claims 1-3 wherein said tissuesample is blood, tumor tissue in node or bone marrow, formalin-fixedembedded histological material, aspirated cytological material or cellson slides.
 5. The method of any one of claims 1-3 wherein saidseparating is accomplished by electrophoresis or chromatography.
 6. Themethod of claim 5 wherein said electrophoresis is agarose orpolyacrylamide gel electrophoresis.
 7. The method of any one of claims1-3 wherein said detection is accomplished by non-isotopic means,radioisotope incorporation, or Southern transfer and hybridization to aninternal probe.
 8. A method according to claim 7, wherein said detectionby non-isotopic means comprises ethidium bromide staining.
 9. The methodof claim 1 or 3 wherein said primers specific for said immunoglobulingene segments are selected from primers to the framework portion of theV regions, primers to parts of the D region, primers to parts of the Jregion, and primers to parts of the C region of the immunoglobulin gene.10. The method of claim 2 or 3 wherein said primers for T-receptor genesegments are selected from primers to the conserved V regions, primersto parts of the D region, primers to parts of the J regions and primersto parts of the C region of the T-receptor gene.
 11. The method of claim1 or 3 wherein said primers specific for said immunoglobulin genesegment are consensus primers for the immunoglobulin gene segment. 12.The method of claim 1 or 3 wherein one of said pair of primers specificfor said immunoglobulin gene segments is a primer to the V region of theimmunoglobulin gene and the other of said primers is a primer to the Jregion of the immunoglobulin gene.
 13. The method of claim 12 whereinsaid primer to the V region of the immunoglobulin gene is FR3A and saidprimer to the J region of the immunoglobulin gene is LJ_(H) ⁻.
 14. Themethod of claim 2 or 3 wherein said primers specific for said T-receptorgene segment are consensus primers for the T-receptor gene segment. 15.The method of claim 2 or 3 wherein one of said pair of primers specificfor said T-receptor gene segments is a primer to the V region of theT-receptor gene and the other of said primers is a primer to the Jregion of the T-receptor gene.
 16. The method of claim 1 or 3 whereinone of said pair of primers specific for said immunoglobulin genesegment is selected from the group consisting of:ELJ_(H) ⁻ : 5'TGAGGAGACGGTGACCAGGGTNCCTTGGCCCCAG 3'; LJ_(H) ⁻ : 5'TGAGGAGACGGTGACC 3';and VLJ_(H) ⁻ : 5' GTGACCAGGGTNCCTTGGCCCCAG 3',and the other of saidprimers is selected from the group consisting of: FR1BR: 5'CT[C/G][T/A]CCTG[T/C][G/A]CAG[T/C]C TCTGG 3'; FR1BS: 5'CTCTCCTGTGCAGCCTCTGG 3'; FR2A: 5'TGG[A/G]TCCG[C/A]CAG[G/C]C[T/C]]T/C]CNGG 3'; FR3A: 5'ACACGGC[C/T][G/C]TGTATTACTGT 3'; and FR3A-: 5'ACAGTAATACA[G/C][A/G]GCCGTGT 3'wherein N represents any nucleotide base.17. The method of claim 2 or 3 wherein said primers specific for saidT-receptor gene segment are selected from the group consisting of:JAT1:5' TTCCAAAGATCAG(T/C)TTG 3'; JAT2: 5' A(A/G)CCTGGT(T/C)CCT(T/G)T/G)TCAAA3'; 3VTA: 5' GA(C/T)TCAGC(T/C)GTGTAC(T/C)(T/A)(C/T)TG 3'; 5VTA1: 5'(A/T)AC(C/T)T(A/C)(C/T)TCTGGTA(C/T)AA(A/G)CA 3'; CTA: 5'GAATAGGCAGACAGACTTGT 3'; PTA: 5' ACTGGATTTAGAGTCTCT 3'; JBT1: 5'CGGGT(G/C)CCT(T/G)GCCC(G/A)AA 3'; JBT2: 5'AGCACGGTGAGCC(G/T)(G/T)GT(G/C)CC 3'; 3VTB1: 5'GAC(T/A)(C/G)A(G/A)(G/C)CGTGTATCT(C/T)TG 3'; 3VTB2: 5' CAGACATCTCTGTAC(C/T)TCTGTGCC 3'; 5VTB1: 5'C(A/T)(C/A)(G/C)(A/C)T(T/G)T(A/T)(T/C)TGGTACCG ACAG 3'; CTB: 5'TTTGGGTGTGGGAGATCTCTGC 3'; PTB: 5' CTTCTGATGGCTCAAACA 3'; JCT12: 5'AAGTGTTGTTCCACTGCCAAA 3'; 3VTG: 5' GTCTATTACTGTGCCACCTGG 3'; CTG: 5'TCTGGAGCTTTGTTTCAGCAA 3'; and PTG: 5' GGAAGAAAAATAGTGGGC 3'.
 18. Themethod of claim 2 or 3 wherein one of said pair of primers specific forsaid T-receptor gene segment is selected from the group consistingof:TCRVG2PST 5' CTTCCTGCAGATGACTCCTACAACTCCAAGGTTG 3'; TCRVG3PST 5'CTTCCTGCAGATGACGTCTCCACCGCAAGGGATG 3'; TCRVG4PST 5'CTTCCTGCAGATGACTCCTACACCTCCAGCGTTG 3'; TCRVG5PST 5'TTCCTGCAGATGACGTCTCCAACTCAAAGGATG 3'; TCRVG8PST 5'CTTCCTGCAGATGACTCCTACAACTCCAGGGTTC 3'; TCRVG9PST 5'GG(A/G/C/T)ACTGCAGGAAAGGAATCTGGCATTCCG 3'; TC4VG10PST 5'CTCTGCAGAATCCGCAGCTCGACGCAGCA 3'; TC4VG11PST 5'CACTGCAGGCTCAAGATTGCTCAGGTGGG 3'; and TC4VG12PST 5'ACTCTGCAGCCTCTTGGGCACTGCTCTAAA 3'and the other of said pair of primersspecific for said T-receptor gene segment is selected from the groupconsisting of: JGT₁₂ : 5' AAGTGTTGTTCCACTGCCAAA 3'; JGT₃ : 5'AGTTACTATGAGC(T/C)TAGTCCC 3'; and JGT₄ : 5' TGTAATGATAAGCTTTGTTCC 3'.19. A kit for the detection of lymphoid leukemia or lymphoma comprisinga first container containing at least one pair of primers specific forimmunoglobulin gene segments wherein said immunoglobulin gene segmentscomprise at least one of a V-D, D-J and J-C junction, and a secondcontainer containing reagents for a polymerase chain reaction.
 20. Thekit of claim 19 wherein one of said pair of primers is selected from thegroup consisting of:ELJ_(H) ⁻ : 5' TGAGGAGACGGTGACCAGGGTNCCTTGGCCCCAG3'; LJ_(H) ⁻ : 5' TGAGGAGACGGTGACC 3'; and VLJ_(H) ⁻ : 5'GTGACCAGGGTNCCTTGGCCCCAG 3',and the other of said primers is selectedfrom the group consisting of: FR1BR: 5'CT[C/T][T/A]CCTG[T/C][G/A]CAG[T/C]C TCTGG 3'; FR1BS: 5'CTCTCCTGTGCAGCCTCTGG 3'; FR2A: 5'TGG[A/G]TCCG[C/A]CAG[G/C]C[T/C][T/C]CNGG 3'; FR3A: 5'ACACGGC[C/T][G/C]TGTATTACTGT 3'; and FR3A-: 5'ACAGTAATACA[G/C][A/G]GCCGTGT 3'wherein N represents any nucleotide base.21. A kit for the detection of lymphoid leukemia or lymphoma comprisinga first container containing at least one pair of primers specific forT-receptor gene segments wherein said T-receptor gene segments compriseat least one of a V-D, D-J and J-C junction, and a second containercontaining reagents for a polymerase chain reaction.
 22. The kit ofclaim 21 wherein one of said pair of primers is selected from the groupconsisting ofTCRVG2PST 5' CTTCCTGCAGATGACTCCTACAACTCCAAGGTTG 3';TCRVG3PST 5' CTTCCTGCAGATGACGTCTCCACCGCAAGGGATG 3'; TCRVG4PST 5'CTTCTGCAGATGACTCCTACACCTCCAGCGTTG 3'; TCRVG5PST 5'TTCCTGCAGATGACGTCTCCAACTCAAAGGATG 3'; TCRVG8PST 5'CTTCCTGCAGATGACTCCTACAACTCCAGGGTTC 3'; TCRVG9PST 5'GG(A/G/C/T)ACTGCAGGAAAGGAATCTGGCATTCCG 3'; TC4VG10PST 5'CTCTGCAGAATCCGCAGCTCGACGCAGCA 3'; TC4VG11PST 5'CACTGCAGGCTCAAGATTGCTCAGGTGGG 3'; and TCRVG12PST 5'ACTCTGCAGCCTCTTGGGCACTGCTCTAAA 3',and the other of said pair of primersis selected from the group consisting of: JGT₁₂ : 5'AAGTGTTGTTCCACTGCCAAA 3'; JGT₃ : 5' AGTTACTATGAGC(T/C)TAGTCCC 3'; andJGT₄ : 5' TGTAATGATAAGCTTTGTTCC 3'.
 23. A kit for the detection oflymphoid leukemia or lymphoma comprising a first container containing atleast one pair of primers specific for immunoglobulin gene segments andat least one pair of primers specific for T-receptor gene segmentswherein said immunoglobulin gene segments and said T-receptor genesegments comprise at least one of a V-D, D-J and J-C junction, and asecond container containing reagents for a polymerase chain reaction.24. The kit of claim 19 or 23 wherein one of said pair of primersspecific for said immunoglobulin gene segment is a primer to the Vregion of the immunoglobulin gene and the second of said pair of primersis a primer to the J region of the immunoglobulin gene.
 25. The kit ofclaim 24 wherein said primer to the V region of the immunoglobulin geneis FR3A and said primer to the J region of the immunoglobulin gene isLJ_(H) ⁻.
 26. The kit of claim 21 or 23 wherein one of said pair ofprimers specific for said T-receptor gene segment is a primer to the Vregion of the T-receptor gene and the second of said pair of primers isa primer to the J region of the T-receptor gene.
 27. The kit of claim 23wherein one of said primers specific for said immunoglobulin genesegment is selected from the group consisting of:ELJ_(H) ⁻ : 5'TGAGGAGACGGTGACCAGGGTNCCTTGGCCCCAG 3'; LJ_(H) ⁻ : 5' TGAGGAGACGGTGACC3'; and VLJ_(H) ⁻ : 5' GTGACCAGGGTNCCTTGGCCCCAG 3',and the other of saidprimers specific for said immunoglobulin gene segment is selected fromthe group consisting of: FR1BR: 5' CT[C/G][T/A]CCTG[T/C][G/A]CAG[T/C]CTCTGG 3'; FR1BS: 5' CTCTCCTGTGCAGCCTCTGG 3'; FR2A: 5'TGG[A/G]TCCG[C/A]CAG[G/C]C[T/C][T/C]CNGG 3'; FR3A: 5'ACACGGC[C/T][G/C]TGTATTACTGT 3'; and FR3A-: 5'ACAGTAATACA[G/C][A/G]GCCGTGT 3'and one of said primers specific for saidT-receptor gene segments is selected from the group consisting of:TCRVG2PST 5' CTTCCTGCAGATGACTCCTACAACTCCAAGGTTG 3'; TCRVG3PST 5'CTTCCTGCAGATGACGTCTCCACCGCAAGGGATG 3'; TCRVG4PST 5'CTTCCTGCAGATGACTCCTACACCTCCAGCGTTG 3'; TCRVG5PST 5'TTCCTGCAGATGACGTCTCCAACTCAAAGGATG 3'; TCRVG8PST 5'CTTCCTGCAGATGACTCCTACAACTCCAGGGTTC 3'; TCRVG9PST 5'GG(A/G/C/T)ACTGCAGGAAAGGAATCTGGCATTCCG 3'; TC4VG10PST 5'CTCTGCAGAATCCGCAGCTCGACGCAGCA 3'; TC4VG11PST 5'CACTGCAGGCTCAAGATTGCTCAGGTGGG 3'; and TCRVG12PST 5'ACTCTGCAGCCTCTTGGGCACTGCTCTAAA 3',and the other of said primers specificfor said T-receptor gene segment is selected from the group consistingof: JGT₁₂ : 5' AAGTGTTGTTCCACTGCCAAA 3'; JGT₃ : 5'AGTTACTATGAGC(T/C)TAGTCCC 3'; and JGT₄ : 5' TGTAATGATAAGCTTTGTTCC 3'.